PROJECT SUMMARY/ABSTRACT Malaria is a significant contributor to global human mortality and accounts for an estimated 445,000 deaths and 216 million cases per year. The vast majority of malaria deaths result from Plasmodium falciparum infection. Resistance to available antimalarials continues to emerge including currently used artemisinin-based combination therapies. New therapeutic targets for treatment of malaria are greatly needed, and target-based drug development relies on our fundamental understanding of essential parasite biology. P. falciparum requires protein prenylation, the C-terminal post-translational lipid modification of proteins, for asexual replication. Our long term goal is to uncover the molecular basis of why prenylation is required by the malaria parasite for survival. We recently identified the complete prenylated proteome of blood-stage P. falciparum malaria parasites. Our preliminary studies revealed that PfHsp40 (PF3D7_1437900), a heat shock protein, is robustly prenylated. Heat shock proteins are necessary for protein folding and stabilization and their expression is upregulated under different cellular stresses including heat shock. P. falciparum experiences heat shock throughout its lifecycle and it is not surprising then that roughly 2% of the P. falciparum genome is dedicated to molecular chaperones such as heat shock proteins. The objective of this proposal is to establish the biological role and regulation of PfHsp40. The proposed studies will answer several fundamental questions including: What is the role of PfHsp40 in P. falciparum? Does prenylation influence the role of PfHsp40 in the parasite? Supported by strong preliminary data that indicate that this strategy will be successful, the objective will be met through two specific aims: 1) Determine the importance of PfHsp40 in asexual replication of P. falciparum; and 2) Determine the mechanism by which protein prenylation controls the biological role of PfHsp40. This approach is innovative, since it uses a combination of state-of-the-art genetic manipulation systems to directly establish the functional aspects of protein prenylation in Plasmodium. The proposed research is significant, because it will illuminate the role of PfHsp40 in the parasite, establish how prenylation influences the biological role of PfHsp40, and identify downstream proteins and pathways that require prenylation in the parasite. Furthermore, because heat shock proteins are found across taxa, the findings are likely to reveal broadly applicable concepts about heat shock protein function.